Assessment of CaMV-mediated gene silencing and integration of CaMV into GM plants with a 35S RNA promoter

Squires, Julie; Stephens, Jennifer; Shoelz, James E.; Palukaitis, Peter

doi:10.1051/ebr:2007043

Cited by 2 publications

(3 citation statements)

References 36 publications

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“…Hypothesizing that the reduction in SC in the T 1 generation, relative to the T 0 transgenic line, might be a result of transgene silencing (which is frequently associated with the 35S promoter [ 63 ]), leading to a reduction in repression of TjYUC6 , we quantified the mRNA levels of TjYUC6 in the T 1 plants ( Figure 8 a–c). The reduction in the phenotype of the T 1 generation was indeed found to be associated with a weaker degree of silencing; both the phenotype and the level of YUC6 knockdown observed in these plants was similar to those of T 0 plant T 0 -41.…”

Section: Resultsmentioning

confidence: 99%

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The S-Gene YUC6 Pleiotropically Determines Male Mating Type and Pollen Size in Heterostylous Turnera (Passifloraceae): A Novel Neofunctionalization of the YUCCA Gene Family

Henning

Shore

McCubbin

2022

Plants

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In heterostylous, self-incompatible Turnera species, a member of the YUCCA gene family, YUC6, resides at the S-locus and has been hypothesized to determine the male mating type. YUCCA gene family members synthesize the auxin, indole-3-acetic acid, via a two-step process involving the TAA gene family. Consequently, it has been speculated that differences in auxin concentration in developing anthers are the biochemical basis underlying the male mating type. Here, we provide empirical evidence that supports this hypothesis. Using a transgenic knockdown approach, we show that YUC6 acts pleiotropically to control both the male physiological mating type and pollen size, but not the filament length dimorphism associated with heterostyly in Turnera. Using qPCR to assess YUC6 expression in different transgenic lines, we demonstrate that the level of YUC6 knockdown correlates with the degree of change observed in the male mating type. Further assessment of YUC6 expression through anther development, in the knockdown lines, suggests that the male mating type is irreversibly determined during a specific developmental window prior to microsporogenesis, which is consistent with the genetically sporophytic nature of this self-incompatibility system. These results represent the first gene controlling male mating type to be characterized in any species with heterostyly.

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Section: Resultsmentioning

confidence: 99%

“…Restriction digests were performed using high-fidelity enzymes (Bam HI and Kpn I), and the CutSmart digest buffer (New England Biolabs, Ipswich, MA, USA). The TjYUC6 fragment was ligated in reverse orientation behind the CaMV 35S promoter in pCHF3, a binary plasmid vector [ 63 ], using T4 ligase (Promega, Madison, WI, USA).…”

Section: Methodsmentioning

confidence: 99%

The S-Gene YUC6 Pleiotropically Determines Male Mating Type and Pollen Size in Heterostylous Turnera (Passifloraceae): A Novel Neofunctionalization of the YUCCA Gene Family

Henning

Shore

McCubbin

2022

Plants

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“…This is further evidence that multiple recombination-excision events occurred in a single plant. Although a number of discreet deletion variants of CaMV were obtained, since CaMV is not seed transmissible (Tompkins, 1937;Squires et al, 2007), these variants would have to be created anew in every generation of plants.…”

Section: Deletions and Insertions In The Sequences Of The Progeny Virmentioning

confidence: 99%

Excision and Episomal Replication of Cauliflower mosaic virus Integrated into a Plant Genome

et al. 2011

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Transgenic Arabidopsis (Arabidopsis thaliana) plants containing a monomeric copy of the cauliflower mosaic virus (CaMV) genome exhibited the generation of infectious, episomally replicating virus. The circular viral genome had been split within the nonessential gene II for integration into the Arabidopsis genome by Agrobacterium tumefaciens-mediated transformation. Transgenic plants were assessed for episomal infections at flowering, seed set, and/or senescence. The infections were confirmed by western blot for the CaMV P6 and P4 proteins, electron microscopy for the presence of icosahedral virions, and through polymerase chain reaction across the recombination junction. By the end of the test period, a majority of the transgenic Arabidopsis plants had developed episomal infections. The episomal form of the virus was infectious to nontransgenic plants, indicating that no essential functions were lost after release from the Arabidopsis chromosome. An analysis of the viral genomes recovered from either transgenic Arabidopsis or nontransgenic turnip (Brassica rapa var rapa) revealed that the viruses contained deletions within gene II, and in some cases, the deletions extended to the beginning of gene III. In addition, many of the progeny viruses contained small regions of nonviral sequence derived from the flanking transformation vector. The nature of the nucleotide sequences at the recombination junctions in the circular progeny virus indicated that most were generated by nonhomologous recombination during the excision event. The release of the CaMV viral genomes from an integrated copy was not dependent upon the application of environmental stresses but occurred with greater frequency with either age or the late stages of plant maturation.

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Assessment of CaMV-mediated gene silencing and integration of CaMV into GM plants with a 35S RNA promoter

Cited by 2 publications

References 36 publications

The S-Gene YUC6 Pleiotropically Determines Male Mating Type and Pollen Size in Heterostylous Turnera (Passifloraceae): A Novel Neofunctionalization of the YUCCA Gene Family

The S-Gene YUC6 Pleiotropically Determines Male Mating Type and Pollen Size in Heterostylous Turnera (Passifloraceae): A Novel Neofunctionalization of the YUCCA Gene Family

Excision and Episomal Replication of Cauliflower mosaic virus Integrated into a Plant Genome

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